Zhongzhi International Institute of Agricultural Biosciences, Shunde Graduate School, Research Center of Biology and Agriculture, University of Science and Technology Beijing (USTB), Beijing 100024, China.
Beijing Engineering Laboratory of Main Crop Bio-Tech Breeding, Beijing International Science and Technology Cooperation Base of Bio-Tech Breeding, Beijing Solidwill Sci-Tech Co. Ltd., Beijing 100192, China.
Int J Mol Sci. 2021 Jul 24;22(15):7916. doi: 10.3390/ijms22157916.
The function and regulation of lipid metabolic genes are essential for plant male reproduction. However, expression regulation of lipid metabolic genic male sterility (GMS) genes by noncoding RNAs is largely unclear. Here, we systematically predicted the microRNA regulators of 34 maize white brown complex members in ATP-binding cassette transporter G subfamily (WBC/ABCG) genes using transcriptome analysis. Results indicate that the transcript was predicted to be targeted by zma-miR164h-5p, and their expression levels were negatively correlated in maize B73 and Oh43 genetic backgrounds based on both transcriptome data and qRT-PCR experiments. CRISPR/Cas9-induced gene mutagenesis was performed on and another lipid metabolic gene, . DNA sequencing, phenotypic, and cytological observations demonstrated that both and are GMS genes in maize. Notably, ZmABCG26 proteins are localized in the endoplasmic reticulum (ER), chloroplast/plastid, and plasma membrane. Furthermore, ZmFAR1 shows catalytic activities to three CoA substrates in vitro with the activity order of C12:0-CoA > C16:0-CoA > C18:0-CoA, and its four key amino acid sites were critical to its catalytic activities. Lipidomics analysis revealed decreased cutin amounts and increased wax contents in anthers of both and GMS mutants. A more detailed analysis exhibited differential changes in 54 monomer contents between wild type and mutants, as well as between and . These findings will promote a deeper understanding of miRNA-regulated lipid metabolic genes and the functional diversity of lipid metabolic genes, contributing to lipid biosynthesis in maize anthers. Additionally, cosegregating molecular markers for and were developed to facilitate the breeding of male sterile lines.
脂质代谢基因的功能和调控对于植物雄性生殖至关重要。然而，非编码 RNA 对脂质代谢基因雄性不育（GMS）基因的表达调控在很大程度上尚不清楚。在这里，我们使用转录组分析系统地预测了玉米 ABCG 亚家族 ATP 结合盒转运蛋白 WBC/ABCG 基因中的 34 个玉米白棕色复合体成员的 microRNA 调节剂。结果表明，基于转录组数据和 qRT-PCR 实验，预测 转录本被 zma-miR164h-5p 靶向，并且它们在玉米 B73 和 Oh43 遗传背景下的表达水平呈负相关。CRISPR/Cas9 诱导的基因诱变在 和另一个脂质代谢基因 上进行。DNA 测序、表型和细胞学观察表明， 和 都是玉米的 GMS 基因。值得注意的是，ZmABCG26 蛋白定位于内质网（ER）、叶绿体/质体和质膜。此外，ZmFAR1 在体外对三种 CoA 底物表现出催化活性，活性顺序为 C12:0-CoA > C16:0-CoA > C18:0-CoA，其四个关键氨基酸位点对其催化活性至关重要。脂质组学分析显示， 和 GMS 突变体的花药中角质层含量减少，蜡质含量增加。更详细的分析显示，在野生型和突变体之间，以及 和 之间，54 种单体含量存在差异变化。这些发现将促进对 miRNA 调控的脂质代谢基因和脂质代谢基因功能多样性的深入理解，有助于玉米花药中的脂质生物合成。此外，还开发了用于 和 的共分离分子标记，以促进不育系的选育。
